Persistent B19 Infection Research Articles

Parvovirus B19: A Potential Cause of Refractory Leukopenia in Lung Transplant Recipients

<h3>Introduction</h3> Leukopenia is common among lung transplant (LT) recipients and medications are the most common offending agents, including valganciclovir (VG), mycophenolate mofetil (MMF), and trimethoprim-sulfamethoxazole (TMS). If leukopenia persists despite stopping culprit medications, patients are typically referred for a bone marrow biopsy (BMB) and parvovirus B19 (B19) testing. In immunocompromised hosts, persistent B19 infection most commonly manifests as pure red cell aplasia and chronic anemia. We report unusual cases of refractory leukopenia in 2 LTR who were found to have B19 on BMB. <h3>Case Report</h3> A 68-year-old woman had bilateral LT (BLT) in February 2020 for idiopathic pulmonary fibrosis (IPF). She had basiliximab induction and our standard immunosuppressive and prophylactic regimen: MMF, VG, and TMS. Four months after BLT she became neutropenic, but not lymphopenic, anemic, or thrombocytopenic. MMF, VG, and TMS were stopped, but she continued to require recurrent filgrastim as her absolute neutrophil count was <500 cells/µL. BMB revealed a hypocellular-for-age bone marrow with diminished, but maturing trilineage hematopoiesis, and PCR detected B19. She was prescribed IVIG, but it was held due to rapidly progressive renal dysfunction. To date, she remains off MMF, VG, and TMS, with letermovir, acyclovir, and pentamidine replacing the latter 2 medications. She is still leukopenic and now also mildly anemic. A 62-year-old man had BLT in August 2020 for IPF. He had basiliximab induction and our standard immunosuppressive and prophylactic regimen: MMF, VG, and TMS. Two months after BLT he became leukopenic and mildly anemic, but not thrombocytopenic. MMF, VG, and TMS were stopped, but he continued to require recurrent filgrastim. BMB revealed a hypocellular-for-age bone marrow (10-15%), and PCR detected B19. He was prescribed IVIG, but IVIG infusions are yet to begin. To date, he remains off of MMF, VG, and TMS, with letermovir, acyclovir, and atovaquone replacing the latter 2 medications. He remains leukopenic, but not anemic or thrombocytopenic. <h3>Summary</h3> We report unusual cases of refractory leukopenia, without significant anemia, in 2 LT recipients with B19 identified on BMB. We suspect that B19 is the culprit as stopping offending medications has been ineffective and hematologic workup has been unrevealing.

Pure Red Cell Aplasia Secondary to Parvovirus B19 Infection in HIV Positive Patients. Clinical and Seroepidemiological Description and Observations in A Third Hospital Level In Colombia

Objective: Describe the clinical and seroepidemiological characteristics and the impact on morbidity and mortality and health costs of patients infected with HIV and red cell aplasia and positive serology for Parvovirus B 19. Methodology: Descriptive and retrospective study, of the clinical and seroepidemiological characteristics of cases of aplasia of the red series without affecting hematimetric indices in HIV positive patients with positive serology to Parvovirus B19, were descripted in an hospitalization service in the period from April 2016 to April 2017. Serum samples from the 16 patients enrolled for the study were analyzed by IgG and IgM type antibodies using the enzyme immunoassay technique. Other causes of anemia, such as: iron-deficiency anemia, invasive fungal infection were ruled out. Results: A total of 16 cases were documented, 11 men (69%) and 5 women (31%) with an average age of 40.7 and 44.4 years respectively, 5 of the 16 patients (31.25%) had positive IgM levels (distributed among 3 men and 2 women) without IgG activity documented in the same sample and 4 of these 5 patients were without treatment for their HIV (80%), none of the patients received regular treatment for HIV. The average in grams per deciliter of hemoglobin and hematocrit at the time of sampling was 8.92 g / dl and 28.6 g / dl respectively, the average of CD4 cell count was 115 for the male population and 187.2 for the female population, the average viral load was 515,483 and 369,390 copies / ml for men and women respectively. No patient received adjuvant treatment directed at the pure red anemia demonstrated. Conclusion: Persistent parvovirus B19 infections in human immunodeficiency virus type 1 infected patients have been reported. The two viruses could share common target cells. The co-infection by Parvovirus B19 is associated with longer days of hospital stay, costs and complications such as nosocomial infections and need of poly transfusions.

Peripheral Blood Mononuclear Cells’ Proliferative Response to Human Parvovirus B19 Antigens in Patients with Rheumatoid Arthritis / Reimatoīdā Artrīta Slimnieku Perifēro Asiņu Mononukleāro Šūnu Proliferativā Atbilde Uz Cilvēka Parvovīrusa B19 Antigēniem

Abstract This study aimed to determine peripheral blood mononuclear cells’ (PBMC) proliferative response to parvovirus B19 (B19) antigens in rheumatoid arthritis (RA) patients and possible changes in proliferative response due to chemotherapy. Serum and blood samples of 52 RA patients and 25 sex and age matched healthy individuals were examined for the presence of anti-B19 IgG and IgM class antibodies and virus specific DNA sequence by the recomLine B19 test and nested polymerase chain reaction, respectively. The PBMC proliferative activity was estimated on the 3rd and 6th day of PBMC cultivation in the presence of virus and B19 VP1/VP2 peptide, using thymidine incorporation assay. On the 3rd day, PBMC response to B19 antigens was detected in 74.1% RA patients with active, in 44.8% - with remote and in 40% - with latent stage of persistent B19 infection, while in the control group the response was observed only in two individuals with active viral infection. On the 6th day, the response was found in 50% RA patients with active, 68.9% - with remote and in 80% - with latent stage of latent persistent infection as well as in 41.1% remotely infected control individuals. The highest PBMC mean stimulation indices were detected in the RA patients and control persons with active infection as well as in RA patients with latent stage of persistent viral infection. On the 3rd and 6th day, strong proliferative response was significantly more frequently observed in RA patients not receiving methotrexate treatment, compared to the patients receiving methotrexate treatment in different combinations with other drugs. RA patients had more frequent and faster response to B19 antigens than apparently healthy persons.

No evidence of persistent parvovirus B19 viremia among Iranian patients with HIV after a 1-year follow-up.

Recent studies have demonstrated that, in common with other latent viruses, parvovirus B19 infection can be controlled by the host immune response but may persist in some places such as the bone marrow. Persistent B19 infection has been found in both immunocompetent and immunocompromised individuals, such as patients infected with human immunodeficiency virus (HIV). However, there is limited data regarding long-term B19 viremia in HIV patients. In this study, we investigated virological and hematological findings, and also the clinical outcome, of seven cases of HIV/B19 coinfection (confirmed by PCR) after one year. These cases were provided from a previous study on patients with HIV infection that found B19 DNA in 13 cases. Seven of these 13 patients were available after 1 year, and we retested them for B19 viremia and B19-specific antibodies. B19 IgG was tested by ELISA, and B19 DNA was assessed by nested PCR. Anemia was not observed in these cases. All subjects had cleared viremia, but B19 IgG seroconversion occurred in two cases. No significant changes in CD4 and hemoglobin occurred. The results of this study indicate that B19 infection in HIV patients is a subtle infection and that B19 viremia is not a long-term event.

Serological and molecular detection of parvovirus b19 in children with acute lymphoblastic leukemia.

Background: Human parvovirus B19 (B19V) is the only human pathogenic parvovirus, It is highly erythrotropic and preferentially replicates in erythroid progenitor cells (EPCs). Recently the effects of B19 infection have been well studied in children with acute lymphoblastic leukemia (ALL).&#x0D; Objective: To detect parvovirus B19 among children with ALL.&#x0D; Methods: A cross sectional study involved forty five patients with ALL (21 patients were newly diagnosed ALL and 24 children who underwent chemotherapy) who were attending department of oncology in Children's Welfare Teaching Hospital in Baghdad Medical City between December 2012 and April 2013, compared to forty five of apparently healthy children as a control group, age and sex were matched. A serological and molecular detection of parvovirus B19 were done using enzyme linked immunosorbant assay (ELISA) and Real-time PCR.&#x0D; Results: B19-IgM, IgG and DNA were detected in study cases as (15.6%, 40% and 13.3%) compared to control group (4.4%, 13 % and 0%) respectively. B19-IgG and B19-IgM were detected in newly diagnosed cases in (41.7% and 19.05%) as well as in (38.1% and 12. 5%) of cases on chemotherapy respectively, all cases with B19-DNA signals were underwent chemotherapy (25%) with mean viral load of (36x104 copies/ml).&#x0D; Conclusion: children who suffering from ALL are at increased risk of B19 infection. Notable percentage of persistent B19 infection was recorded among children with ALL who are receiving ongoing treatment.

SAT0071 Dependence of rheumatoid arthritis disease aggressivity and activity on the presence of parvovirus B19 infection markers

BackgroundThe role of parvovirus B19 (B19) infection as an initiator of the classic erosive rheumatoid arthritis (RA) is being constantly debated. The main argument against etiological association of B19 with...

Human immunodeficiency virus/human parvovirus B19 co-infection in blood donors and AIDS patients in Sichuan, China.

Human parvovirus B19 (B19) is a common pathogen which causes a variety of diseases. Persistent B19 infection is related to the degree of host immunodeficiency in patients with human immunodeficiency virus (HIV) infection. However, the existence, loading, virus evolution and distribution of B19 in Chinese HIV-positive patients have not been determined. We investigated 573 HIV-positive blood donors and AIDS patients in Sichuan, China in the last two decades. Bl9-specific serology and quantitative polymerase chain reaction were used to determine the prevalence of B19/HIV co-infection. Viral genome fragments were subjected to phylogeny and haplotype analysis. B19 genomic DNA was found in 26 of 573 (4.5%) HIV-positive individuals, a higher prevalence than in blood donors. DNA levels ranged from 5.3×10(2)-1.1×10(5) copies/mL. The seroprevalence of IgG was significantly lower in HIV-positive samples than in HIV-negative blood donors, indicating deficient production of B19-specific IgG in the former. The B19 isolates were genotype-1 subtype B19-1A which formed a monophyletic group; seven distinct haplotypes were discovered with 60% of the B19/HIV co-infected variants sharing one central haplotype. This study on the prevalence, phylogeny and distribution of human parvovirus B19 in Sichuan, China, demonstrates the persistence of B19 in the circulation of both immunocompetent and immunocompromised subjects, with implications for blood safety.

Anemia as a Complication of Parvovirus B19 Infection in Renal Transplant Recipients

The frequency of B19 infection in renal transplant donors and recipients was studied to determine the significance of active viral infection in the development of anemia. Serum, plasma, and peripheral blood leukocyte samples of 47 renal transplant donors, 38 recipients with anemia (Group 1), and 25 without anemia (Group 2) after renal transplantation were evaluated for the presence of anti-B19 specific antibodies (ELISA) and B19 DNA (nPCR). Active persistent B19 infection after renal transplantation was detected in 12 of the 38 in the Group 1 (10 had reactivation and 2 primary infection), and none of the recipients in the Group 2 had it. Of the 12 recipients in the Group 1, 10 were seropositive and 2 seronegative before renal transplantation; 10 received the transplants from the seropositive and 2 from seronegative donors. rHuEPO therapy-resistant severe anemia was detected only in the recipients with active B19 infection after renal transplantation in the Group 1 (7/12). The logistic regression analysis revealed a significant relationship between active B19 infection and severe anemia (OR, 0.039; 95% CI, 0.006-0.257; P=0.001). Active B19 infection was documented only in the anemic recipients and could be associated with the development of severe anemia after renal transplantation. This allows us to recommend concurrent screening for viral DNA in plasma and detection of anti-B19 IgM class antibodies. To find the association between B19 infection and the development of anemia, further investigations are necessary.

Parvovirus B19 Genotype Specific Amino Acid Substitution in NS1 Reduces the Protein's Cytotoxicity in Culture

A clinical association between idiopathic liver disease and parvovirus B19 infection has been observed. Fulminant liver failure, not associated with other liver-tropic viruses, has been attributed to B19 in numerous reports, suggesting a possible role for B19 components in the extensive hepatocyte cytotoxicity observed in this condition. A recent report by Abe and colleagues (Int J Med Sci. 2007;4:105-9) demonstrated a link between persistent parvovirus B19 genotype I and III infection and fulminant liver failure. The genetic analysis of isolates obtained from these patients demonstrated a conservation of key amino acids in the nonstructural protein 1 (NS1) of the disease-associated genotypes. In this report we examine a conserved residue identified by Abe and colleagues and show that substitution of isoleucine 181 for methionine, as occurs in B19 genotype II, results in the reduction of B19 NS1-induced cytotoxicity of liver cells. Our results support the hypothesis that in the setting of persistent B19 infection, direct B19 NS1-induced cytotoxicity may play a role in idiopathic fulminant liver failure.

Different patterns of disease manifestations of parvovirus B19-associated reactive juvenile arthritis and the induction of antiphospholipid-antibodies

Children with rheumatic oligo- and polyarthritis frequently establish persistent parvovirus B19 infections, which may be associated with the production of antiphospholipid antibodies. Reported in this paper are the data of five girls with polyarticular rheumatic diseases of different types and persistent parvovirus B19 infection associated in four cases with the presence of antibodies against phospholipids. Clinical parameters, virus load, and antiphospholipid-IgG levels were determined during an observation period up to 92 months. In two patients, erythema infectiosum preceded the development of arthritis and B19 viremia persisted. Two other girls showed antibodies against parvoviral structural proteins at time of the manifestation of the rheumatic disease. Subsequent samples also revealed persistent B19 infection. In the fifth patient, parvovirus B19-specific IgG antibodies were detected for the first time after 120 months of progressing disease at an age of 11 1/2 years. Five years later, quantitative polymerase chain reaction (PCR) revealed viral DNA. In a synovial tissue specimen subsequently obtained, parvovirus B19 structural proteins could be detected by immunohistochemistry. Three of five patients recovered completely without severe sequels. One patient is in remission under immunosuppressive therapy. The fifth patient suffers from progressive erosions despite intensive therapeutical efforts. In consequence, parvovirus B 19 should generally be taken into consideration as a trigger of various forms of juvenile arthritis and persistence of infection should be evaluated.

Cytokine responses in acute and persistent human parvovirus B19 infection

The aim of this study was to characterize the proinflammatory and T helper (Th)1/Th2 cytokine responses during acute parvovirus B19 (B19) infection and determine whether an imbalance of the Th1/Th2 cytokine pattern is related to persistent B19 infection. Cytokines were quantified by multiplex beads immunoassay in serum from B19-infected patients and controls. The cytokine responses were correlated with B19 serology, quantitative B19 DNA levels and clinical symptoms. In addition to a proinflammatory response, elevated levels of the Th1 type of cytokines interleukin (IL)-2, IL-12 and IL-15 were evident at time of the initial peak of B19 viral load in a few patients during acute infection. This pattern was seen in the absence of an interferon (IFN)-gamma response. During follow-up (20-130 weeks post-acute infection) some of these patients had a sustained Th1 cytokine response. The Th1 cytokine response correlated with the previously identified sustained CD8+ T cell response and viraemia. A cross-sectional study on patients with persistent B19 infection showed no apparent imbalance of their cytokine pattern, except for an elevated level of IFN-gamma response. No general immunodeficiency was diagnosed as an explanation for the viral persistence in this later group. Neither the acutely infected nor the persistently infected patients demonstrated a Th2 cytokine response. In conclusion, the acutely infected patients demonstrated a sustained Th1 cytokine response whereas the persistently infected patients did not exhibit an apparent imbalance of their cytokine pattern except for an elevated IFN-gamma response.

Genetic drift of parvovirus B19 is found in AIDS patients with persistent B19 infection

It is generally thought that parvovirus B19 is stable genetically. Consistently, genetic drift has not been found in patients with persistent B19 infection. In this report, longitudinal genetic changes in NS1 and VP1 gene of B19 isolates from three AIDS patients with persistent B19 infection were studied. One of the three patients was not treated with highly active anti-retroviral therapy (HAART). B19 viral DNA from these patients was amplified by polymerase chain reaction (PCR) and then sequenced directly. A single genetic change was found in the B19 isolate obtained from the patient not treated with HAART on Day 10 after intravenous immunoglobulin (IVIG) treatment. The nucleotide sequences of B19 isolated from this patient, then remained unchanged over a period of 11 months. Analysis of NS1 clones derived from his longitudinal viral isolates showed the existence of quasi-species but genetic drift was not found. One of the other two patients treated with HAART experienced treatment failure; he was later treated with mega-HAART. In contrast to the genetic stability of B19 isolates from the patient not treated with HAART, multiple genetic changes were discovered in the viral isolates from the two other patients after HAART and mega-HAART, respectively. Through analysis of B19 clones, the frequency of clones containing these mutations confirmed the genetic drift. Nucleotide substitutions seen in VP2 gene of isolates with genetic drift from both patients were all non-conserved, suggesting that they are positively selected.

Establishment of functional B cell memory against parvovirus B19 capsid proteins may be associated with resolution of persistent infection

Parvovirus B19 (B19) infection can occur during acute lymphoblastic leukemia and persistent viral infection can occur despite intravenous immunoglobulin administration. Here, evidence is presented that resolution of persistent B19 infection in an acute lymphoblastic leukemia patient may be associated with the simultaneous strengthening of antigen-specific B cell memory against the B19 capsid protein VP2 and diminution in the memory response against the B19 non-structural protein 1 (NS1). Determination of antigen-specific B cell memory status may enhance the serological and molecular analyses of persistent B19 infection.

Persistent B19 infection in immunocompetent individuals: implications for transfusion safety

AbstractRecent reports suggested that parvovirus B19 (B19) might persist in immunocompetent individuals such as blood donors, but only cross-sectional data were available. Serial samples from a cohort of multitransfused patients with hemoglobinopathies and a cross-sectional population of pregnant women were tested for B19 markers. Of 76 red cell recipients, 6 (8%) had persistent viral DNA for 1 to 3 or more years, depending on the sensitivity of the genomic amplification assay. All patients also carried B19-specific immunoglobulin G (IgG). In contrast, 0.8% of 500 pregnant women carried both detectable B19 DNA and specific IgG. These results demonstrate that persistence of low levels of B19 DNA suggested by cross-sectional studies is frequent in multitransfused patients and that the virus may remain detectable several years after infection in nonimmunodeficient individuals. (Blood. 2005;106:2890-2895)

CD4+ T‐Cell Responses Against the VP1‐Unique Region in Individuals with Recent and Persistent Parvovirus B19 Infection

To date cellular immune responses against parvovirus B19 (B19) have not been studied extensively. The aim of this study was to examine the T-cell response against the VP1-unique region as the immunodominant part of the viral structural protein VP1 in individuals with different courses of B19 infection. Therefore, a group of 13 parvovirus-positive probands was separated into subgroups characterized for recent or acute, past or persistent infection by means of the presence of specific immunoglobulin (Ig)M and IgG isotypes and of viral DNA in blood and tissue. Transiently transfected B-cells expressing VP1-unique region were used in ELISpot assays to investigate T-cell responses directed against the VP1-unique region in peripheral blood mononuclear cells (PBMC) of individual donors. Significant numbers of interferon-gamma (IFN-gamma) secreting lymphocytes were detectable in PBMC of all individuals with recent, acute or persistent B19 infection, but not in PBMC of donors with past B19 infection and seronegative individuals. A more detailed analysis of IFN-gamma producing cells by intracellular cytokine staining by flow cytometry revealed, that CD4(+) T cells but not CD8(+) cytotoxic lymphocytes (CTL) were the major subpopulation of IFN-gamma producing cells. These data strongly suggest the need of virus protein production for the maintenance of VP1-unique region-specific CD4(+) T-helper cell responses in B19-infected individuals.

Persistent human parvovirus B19 infection in children under maintenance chemotherapy for acute lymphocytic leukemia.

To report on B19 infection management and chemotherapy schedule consequences in five children treated for acute lymphocytic leukemia (ALL). Between May 2001 and February 2002, five patients between 4 and 12 years of age, receiving maintenance chemotherapy for ALL, presented with symptoms suggesting B19 infection (pallor, fatigue, petechiae and pancytopenia in four patients; generalized rash in two patients; acute hepatitis in one patient). Qualitative polymerase chain reaction (PCR) on peripheral blood was used for diagnosis and follow-up of infection; quantitative PCR was used for viral load measurement. Intravenous nonspecific high-dose immunoglobulin therapy was administered until PCR was negative. Qualitative B19 DNA was found in the peripheral blood of all patients, confirming the infection. Viral load at diagnosis ranged from 10 to 10 particles/mL blood. B19 DNA was detectable in four patients at 45, 21, 40, and 44 weeks, respectively. Chemotherapy was delayed in all patients. No clear benefit of intravenous immunoglobulin was noted. Infection with B19 is rarely reported in patients with ALL, but it should be suspected when unexplained pancytopenia occurs during chemotherapy. Persistent B19 infection remains a challenge in the management of patients receiving maintenance chemotherapy for ALL, as no specific therapy such as a specific immunoglobulin or vaccine exists. The role of viral load measurement needs to be established in terms of its use in follow-up and evaluation of the therapeutic response.

Antibody response to B19 parvovirus VP1 and VP2 linear epitopes in patients with haemophilic arthritis.

Parvovirus B19 infection occurs very frequently in patients with haemophilia on account of its transmission with plasma derivatives. In order to achieve a more defined serological pattern for the study of the role of B19 infection in haemophilic arthritis, 53 serum samples from 37 patients with haemophilic arthritis were investigated for the presence of IgG immune response against B19 VP2 and VP1 linear epitopes and VP2 conformational antigen compared to the serological reactivity against B19 NS1 and to the presence of B19 DNA in the synovial membranes. An IgG immune response against VP1 and VP2 linear epitopes was detected by immunoblot assay using recombinant proteins expressed in Escherichia coli. Specific IgG against VP2 and VP1 linear epitopes were present in 84.90 and 92.45% of haemophilic arthritis patients and in 28.0 and 64.0% of the controls (P<0.001) respectively. All 53 sera of the haemophiliacs (100%) and 66.0% of the controls (P<0.001) were IgG positive and IgM negative against VP2 structural epitopes. Specific IgG against VP2 linear epitopes, which are a serological marker of active or very recent B19 infection, proved to be significantly associated with the presence of anti-NS1 antibodies and with the presence of B19 DNA in synovial tissue in patients with haemophilic arthritis. In conclusion, in these patients the presence of B19 IgG anti-VP2 linear epitopes, in absence of IgM anti-VP2 structural antigens, can be a useful serological marker to diagnose active, recent or persistent B19 infection.

Persistent Parvovirus B19 Infection without the Development of Chronic Anemia in HIV‐Infected and ‐Uninfected Children: The Women and Infants Transmission Study

We evaluated the prevalence of persistent parvovirus B19 (B19) infection and associated anemia in human immunodeficiency virus (HIV)-infected and HIV-uninfected children. B19 persistence was defined as B19 DNA detected in specimens collected >16 weeks apart. Of 182 children, 3 HIV-infected children and two HIV-uninfected children had evidence of persistent B19 infection. Of the 5 children, none had evidence of B19-associated anemia. Our data suggest that B19 infections can persist in children without the development of symptomatic anemia.

Intravenous immunoglobulin treatment of four patients with juvenile polyarticular arthritis associated with persistent parvovirus B19 infection and antiphospholipid antibodies

Children with rheumatic oligoarthritis and polyarthritis frequently establish persistent parvovirus B19 infections that may be associated with the production of antiphospholipid antibodies (anti-PL IgG). In this study we analysed the influence of high-dose intravenous immunoglobulin (IVIG) therapy on virus load, on the level of anti-PL IgG and its potential capacity to improve the patients' clinical status. Four juvenile patients with long-lasting polyarticular rheumatic diseases and persistent parvovirus B19 infection, associated in three cases with the presence of antibodies against β2-glycoprotein I (anti-β2GPI IgG), were treated with two cycles of IVIG on five successive days (0.4 g/kg per day). Clinical parameters including scores of disease activity, virus load and anti-PL IgG levels were determined before, during and after treatment. Two patients showed a complete remission that has lasted 15 months. During that period they showed neither clinical nor laboratory signs of inflammation. Viral DNA was not detectable in serum, and a decrease in anti-β2GPI IgG was observed. As assessed by the Childhood Health Assessment Questionnaire and the Health-related Quality of Life Questionnaire for Children, both patients were no longer restricted in their activities of daily living and no impact on the health-related quality of life was observed. In one patient the therapy failed: there was no improvement of symptoms and no decrease in virus load or inflammatory parameters. In the fourth patient, clinical and laboratory parameters did not improve despite a decrease in both viral load and anti-PL IgG. Our results show that the use of IVIG to treat parvovirus B19-triggered polyarticular rheumatic disease of childhood might offer an opportunity to improve this disabling condition.

Relevance of B19 markers in serum samples for a diagnosis of parvovirus B19-correlated diseases.

In order to evaluate the optimal and essential diagnostic test(s) for a correct diagnosis of B19 diseases, 344 consecutive serum samples were tested from 344 patients with clinical suspicion of B19 infection during an epidemic period (early Spring-Autumn 2000). Sera were tested for B19 DNA by a standardized competitive polymerase chain reaction-enzyme-linked immunosorbent assay (PCR-ELISA) and dot-blot hybridization and for specific IgM and IgG by ELISA. Of 344 patients examined, 125 were positive for markers of B19-associated disease: 49 had both B19 DNA and IgM, 50 had B19 DNA without IgM, and 26 had IgM without B19 DNA. After examination of the different patterns of B19 markers as diagnostic tools for B19 infection, IgM determination detected only 60% of B19-documented infections. IgM tests were nevertheless fundamental, as they were the unique diagnostic marker in 20.8% of documented infections (26 of 125 patients), in the diagnosis of recent, but still symptomatic infections when B19 DNA was no longer detectable. The determination of B19 DNA with PCR permitted detection of 79.2% of infections and therefore represented an essential test. PCR was fundamental for the diagnosis of B19 disease, as the unique diagnostic marker in 32% of documented infections (50 of 125 patients), both in acute infections at the onset of symptoms before the appearance of immunological response, and during the course of persistent B19 infections in which IgM had cleared. The contemporaneous determination of B19 DNA by PCR and specific IgM appears to be the most appropriate diagnostic protocol for the correct laboratory diagnosis of B19 infection.